Re-Circulation Loop in CFF/TFF Single Use Path Flow

ABSTRACT

The disclosed subject matter relates to an automated CFF/TFF system that provides the operations of system treatment with a system treatment solution(such as buffer) and filtration of a solution (e.g., a biofluid) can be accomplished in succession without having to re-configure the system (such as re-routing the fluid conduits) that could compromise the integrity of the system. As a result, there will be reduced contamination risk, reduced volume of buffer required and will save time for the filtration process.

BACKGROUND

The present invention relates to an automated crossflow filtration method and system for separating a component of interest from one or more other components in a solution. The invention is of use in the field of purifying bio fluids including protein separations, where specific proteins must be separated and purified from cell lysates and cultures.

Cell culture and bioreactors and various biotechnology processes have generated considerable interest in recent years due to increased interest in genetic engineering and biopharmaceuticals. Cells are cultured to make, for example, proteins, receptors, vaccines, and antibodies that are utilize in patient therapy, research, as well as in various diagnostic techniques.

The separation of such molecules is of increasing commercial interest in the chemical, pharmaceutical and biotech industries, including, for example, the production of biological drugs and diagnostic reagents. In particular, the isolation and purification of proteins is of increasing importance because of advances in proteomics (the study of the function of proteins expressed by the human genome). Some proteins may only be present at very low concentrations within a biological sample. As a result, development of isolation and separation techniques is particularly important, especially when large scale production is involved.

In general, when proteins are produced in cell culture, they are either located intracellularly or secreted into the surrounding culture media. Also, the cell lines that are used are living organisms and must be fed with a growth medium, containing sugars, amino acids, growth factors, etc. Separation and purification of a desired protein from such a complex mixture of nutrients as well as cellular by-products, to a level sufficient for characterization, poses a formidable challenge.

Ultrafiltration membranes are characterized by pore sizes which enable them to retain macromolecules. Such macromolecules may have a molecular weight ranging from about 500 to about 1,000,000 daltons. As such ultrafiltration membranes can be used for concentrating proteins. Ultrafiltration is a low-pressure membrane filtration process which can be used to separate solutes up to 0.1 μm in size. As a result, for example, a solute of molecular size significantly greater than that of the solvent molecule can be removed from the solvent by the application of a hydraulic pressure, which forces only the solvent to flow through a suitable membrane (usually one having a pore size in the range of 0.001 to 0.1 μm). As a result, ultrafiltration is capable of removing bacteria and viruses from a solution.

Crossflow filtration (“CFF” also referred to a “tangential flow filtration” (TFF)) systems can be are used in industry applications, such as, for example, manufacturing process separations, waste treatment plants and water purification systems where they can extend the lifetime of filtration membranes by removing and/or preventing the build-up of contaminants and promote consistency of the filtration process with time.

The most commonly used CFF/TFF membrane processes are microfiltration and ultrafiltration. Such processes may be pressure driven and depend upon the “membrane flux”, defined as the flow volume over time per unit area of membrane, across the microfiltration or ultrafiltration membrane. At low pressures, the transmembrane flux is proportional to pressure. As a result, by varying the transmembrane pressure difference driving force and average pore diameter, a membrane may serve as a selective barrier by permitting certain components of a mixture to pass through while retaining others. This results in two phases, the permeate and retentate phases, each of which is enriched in one or more of the components of the mixture. The retentate stream is recirculated in the flow circuitry and is pumped across the membrane again in a continuous fashion. Such CFF/TFF systems are used to significantly reduce the volume of the sample solution as a permeate stream is withdrawn from the system. So, the sample solution becomes concentrated when the system is driven in a concentration mode.

CFF/TFF systems have the advantage that due to the direction of the flow of the fluid sample, which is essentially parallel to the membrane surface, an automatic sweeping and cleansing takes place so that higher fluxes and higher throughputs can often be attained with such systems in relation to corresponding normal flow filtration systems. Further, a large fraction of sample flows continuously over the membrane surface so that a clogging and fouling is discouraged in such systems. With respect to these and other advantages, CFF/TFF systems are often used in industrial and/or biotechnological processes.

In an automated CFF/TFF system, buffer and other system treatment solutions need to circulate through the filter and other system components for equilibration prior or subsequent to the separation process. Ideally, such circulation and equilibration of buffer and other system treatment solutions is performed by an automated method without the need for manual intervention.

BRIEF DESCRIPTION

In one embodiment, an automated CFF/TFF system configured to utilize flexible tubing is provided. The automated CFF/TFF system comprises a cabinet and an electronic data processing network. The cabinet has sides that define a cabinet interior and a cabinet exterior and includes a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior and a plurality of sensor connectors such that when a sensor is connected thereto the portion of the sensor to which the flexible tubing is connected is positioned on the cabinet exterior. The electronic data processing network is at least partially positioned in the cabinet interior and is connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors. The plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors are proximate to a path for flexible tubing configured to form a fluid circuitry including a reservoir and a CFF/TFF filtration module through which a fluid stream of a solution may be conducted through the automated CFF/TFF system. The fluid circuitry comprises a feed conduit to provide fluid communication between an outlet of the reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module and a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit.

In another embodiment, a method of using an automated CFF/TFF system is provided. The method includes providing an automated CFF/TFF system comprising a cabinet and an electronic data processing network. The cabinet has sides that define a cabinet interior and a cabinet exterior and includes a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior and a plurality of sensor connectors such that when a sensor is connected thereto the portion of the sensor to which the flexible tubing is connected is positioned on the cabinet exterior. The electronic data processing network is at least partially positioned in the cabinet interior and is connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors. The plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors are proximate to a path for flexible tubing configured to form a fluid circuitry including a first reservoir, a second reservoir, a third reservoir and a CFF/TFF filtration module through which a fluid stream of a solution may be conducted through the automated CFF/TFF system. The fluid circuitry comprises a feed conduit to provide fluid communication between an outlet of the reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module, a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit, a second reservoir feed conduit to provide fluid communication between the first reservoir and the second reservoir, a third reservoir feed conduit to provide fluid communication between the first reservoir and the third reservoir, and a drain. The method also includes connecting a sensor to at least one of the plurality of sensor connectors; forming the fluid circuitry using flexible tubing to connect the first reservoir, the second reservoir, the third reservoir, the CFF/TFF filtration module, the plurality of pinch valves, the plurality of pumps and the plurality of sensors; configuring the plurality of pumps and plurality of valves to provide a system treatment solution to the first reservoir from the second reservoir; providing a system treatment solution to the first reservoir from the second reservoir; configuring the plurality of pumps and plurality of valves to circulate the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; circulating the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; configuring the plurality of pumps and plurality of valves to drain the system treatment solution from the automated CFF/TFF system using the drain; draining the system treatment solution from the automated CFF/TFF system using the drain; configuring the plurality of pumps and plurality of valves to provide a solution for filtration to the first reservoir from the third reservoir; providing a solution for filtration to the first reservoir from the third reservoir; configuring the plurality of pumps and plurality of valves to filter the solution for filtration from the first reservoir through the CFF/TFF filtration module; and filtering the solution for filtration from the first reservoir through the CFF/TFF filtration module. In another embodiment, an automated CFF/TFF system is provided. The automated CFF/TFF system comprises a cabinet, an electronic data processing network and flexible tubing. The cabinet has sides that define a cabinet interior and a cabinet exterior and includes a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior, and a plurality of sensor connectors including a sensor is connected to at least one of the plurality of sensor connectors, the portion of the sensor to which the flexible tubing is connected is positioned on the cabinet exterior. The electronic data processing network is at least partially positioned in the cabinet interior and is connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensors. The flexible tubing forms a fluid circuitry including a reservoir and a CFF/TFF filtration module through which a fluid stream of the solution may be conducted through the automated CFF/TFF system and comprises a feed conduit to provide fluid communication between an outlet of the reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module and a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit.

Further suitable embodiments of the invention are described in the dependent claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof.

FIG. 2 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof that shows a filling operation before the filtration process.

FIG. 3 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof shows the filtration process.

FIG.4 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof shows a filling operation of system treatment solution.

FIG. 5 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof shows a circulation operation of system treatment solution.

FIG. 6 illustrates a schematic flow diagram of a CFF/TFF system according to one embodiment thereof shows a draining operation of system treatment solution.

FIG. 7 illustrates a block diagram of an exemplary computing apparatus.

FIG. 8 illustrates a cabinet for the CFF/TFF system of FIG. 1 (outside view). a) Right side panel, b) Frontal panel, c) Left side panel.

FIG. 9 illustrates the FIG. 8 cabinet, with the panels shown from the inside. Dashed lines indicate electrical connections.

DETAILED DESCRIPTION

In an automated cross flow filtration system the buffer preferably re-circulates through the filter for equilibration by an automated method without the need for manual intervention. The flow path should be designed for this purpose in a way that can serve the re-circulation loop for re-circulating the equilibration buffer for filter conditioning or cleaning solution for filter cleaning and storage. A re-circulation loop in a CFF/TFF single use flow path will address the need for buffer re-circulation in an automated method. This method of automated buffer re-circulation will eliminate manual steps, save time for the filtration process and reduce the volume of buffer or other system treatment solution required. One benefit resulting from these advantages is reduced cost of operating the filtration system. Another benefit resulting from these advantages is that the sterility and/or integrity of the fluid system is not compromised as well as risk for compromising the sterility and/or integrity of the fluid system is greatly reduced by eliminating manual interaction for connecting or disconnecting fluid conduits and containers at the outlet and/or inlet connections of the fluid system during the process.

Biofluids as used herein refers to fluids prepared by biological or pharmaceutical methods and may contain biological agents such as cells, molecules (particularly, valuable proteins), suspended particles, media, buffer, carrier, reaction solution, or other liquid component.

System treatment solutions as used herein refers to buffers including equilibration buffers and other pre-filtration system treatment solutions, post filtration system treatment solutions including washing fluid and/or buffers and storage solutions placed into the system during inactivity.

One embodiment of a CFF/TFF system 100 according to the invention, utilizing a microfiltration membrane is shown in FIG. 1, FIG. 8 and FIG. 9. The system may be automated and can be used to separate components present in a solution, such as are commonly found in biological samples including biofluids. For example, depending upon the pore size of the membrane used, cells (such as blood cells) can be washed with buffers prior to lysis to remove contaminants, cellular debris can be separated from soluble materials, and/or proteins can be purified for characterization.

The exemplified embodiment of FIG. 1 includes an automated system 100 that comprises reservoir 102, reservoir 104 and filter 106, for example a CFF/TFF filter or filtration module. The volume of reservoirs 102 and 104 can, for example, range from around 1 liter to around 200 liters in volume and can be scaled to around 2000 liters or more. The reservoirs may be single use or multi-use and can include bags or other vessels composed of suitable material, such as, for example, glass, ceramics or stainless steel and inert plastic material, such as flexible material including flexible plastic material. A means for mixing or agitating the contents of the reservoirs may also be utilized in conjunction with the reservoirs.

The filter 106 may include a microfiltration membrane but it will be understood that, depending upon the nature of the separation to be effected, an ultrafiltration or other membrane could be used. The membrane may be flat or hollow in configuration. A microfiltration membrane may be chosen which has pore sizes such that the component of interest (e.g., a protein) within the solution will pass through the membrane whereas larger components will be retained by it or such that the component of interest (e.g., a protein) within the solution will be retained by the membrane whereas smaller components will pass through it. The material contained therein passing through the membrane is known as the permeate, while the material retained by the membrane is called the retentate.

Suitable membranes may include ultrafiltration, microporous, nanofiltration or reverse osmosis filters formed from polyvinylidene fluoride (PVDF), polysulfone, polyethersulfone, polyarylsulfone, regenerated cellulose, polyamide, polypropylene, polyethylene, polytetrafluoroethylene, cellulose acetate, polyacrylonitrile, vinyl copolymer, polyamides (such as “Nylon 6” or Nylon 66”) polycarbonate, PFA, blends thereof or the like.

The system may also includes a plurality of pumps, valves and sensors (sensors as used herein may also includes meters) for driving, regulating, and acquiring data about a solution as it moves through the system. The system may also include an electronic data processing network capable of receiving, transmitting, processing and recording data related to as well as coordinating the operation of the pumps, valves, sensors and the filter. The sensors may be used to gather information and data about the solution passing through the system at various points therein. Such sensors may monitor physical parameters within the system, such as, for example, pressure, temperature, conductivity, pH, oxygen concentration and ultraviolet light absorption as well as volume flow and/or mass flow measurements.

The system includes a plurality of pumps to drive the flow of solution through the system. Although pumps are preferred, other electronically-controllable means for driving a sample solution through such a system may also be used. Pumps used may include, for example, high-pressure positive displacement (HPPD) pumps, diaphragm pumps, piston pumps, centrifugal, lobe- and peristaltic pumps as well as other pump configurations including, for example, piezoelectric-driven, acoustically-driven, thermopneumatically-driven and electrostatically-driven pumps. Suitable pump flow rates may range from about 10 ml. per minute to about 1000 liters per minute, preferably about 20 ml. per minute to about 400 liters per minute, depending on feed volume and material to be separated with a stipulated time period.

For certain biopharmaceutical applications in which the biofluid under investigation has substantial and significant protein content, forces and circumstances that can lead to the unintended and undesired denaturation of proteins (i.e., the loss of the physical conformation of the protein's polypeptide constituency) should be avoided and/or mitigated. The mechanical shear forces often produced in the operation of certain pumps, particularly at gas/liquid interfaces (cf. e.g., bubbles), have been linked to protein denaturation, and accordingly, should be mitigated and/or avoided in the selection, manufacture, and incorporation of the device.

Valves are positioned along or otherwise functionally proximate the solution path for regulating the flow of the solution therethrough. For example, the valves may be pinch valves that apply a force to flexible tubing passing therethrough to regulate the flow of the solution. The flow of solution through a valve depends upon whether the valve is in an “open” or “closed” state or in some circumstances in an intermediate state, the latter being where the flow of solution is not completely “on” or “off” but in between, thereby regulating the flow rate therethrough.

A conduits system connects reservoir 102, reservoir 104 and filter 106 and so that they are in fluid communication and includes a series of conduits that may also include various valves, pumps and sensors. The filter 106 may include a CFF/TFF filter. There is no particular limitations to the type of conduit used. Potential conduit types include, for example, rigid pipes, flexible tubing, and the channels and passages formed in or intrinsic to the system's other components (e.g., filter, sensors, valves and pumps). Other conduit systems may include “cassettes” that integrate multiple components of the flowpath, for example. For example, tubing networks or fluid conduit networks may be integrated into a single device that may also include valves to regulate the flow through the tubing network, as well as sensors, pumps, filtration modules and other fluid treatment, control or monitoring components. Typically, the plurality of conduits employed in the system may include a mixture of conduit types. Preferably, the majority of the conduits employed are flexible, substantially biologically inert, synthetic polymeric tubing having an internal diameter of 1-50 mm, more preferably 3-32 mm.

In the embodiment of FIG. 1 reservoir 102 is in fluid communication with conduit 108 that is in fluid communication with conduit 110, the latter may include valve 112. A second conduit section includes conduit 114 and valve 116. Conduit 114 may be in fluid communication with another reservoir (not shown) that may be similar to reservoirs 102 and 104. Conduits 110 and 114 are both in fluid communication with conduit 118 that may include pump 120 and valves 122 and 124. Conduit 118 is in fluid communication with conduit 125 that includes valve 126 and with conduit 128 that includes valve 130.

Conduit 125 is in fluid communication with conduit 132 that is connected to retentate outlet 134 of filter 106 and to reservoir 104. Conduit 132 also includes valves 136, 138 and 140 (such as, for example, external tube pinch valves) as well as various sensors including, for example, flow sensor (F) 142, conductivity and temperature sensor (C&T) 144 and pressure retentate sensor (Pr) 146. Conduit (alternatively called a feed conduit) 148 is in fluid communication with reservoir 104 and with conduit 132 via reservoir 104. Conduit 148 is also in fluid communication with feed inlet 150 of filter 106. Conduit 148 may include pump 152, valves 154 and 156 as well as various sensors including, for example, pressure feed sensor (Pf) 158. Valve 160 is also in fluid communication with conduit 148 and a drain/drain line 161 and is connected to a drain/collection/recovery/sample container (not shown).

The embodiment may also include valves and air filters connected thereto such that the valves are in fluid communication with system conduits (not shown). The air filters may be open to the atmosphere or connected to an external integrity tester. Filter 106 also includes permeate outlets 170 and 172 that are both in fluid communication with conduit 174. Conduit 174 is in fluid communication with conduit 176 and further with conduit 186, forming together a permeate conduit. Conduit 176 includes valve 178 as well as various sensors including, for example, flow sensor (F) 180, conductivity and temperature sensor (C&T) 182 and pressure permeate sensor (Pp) 184. Conduits 176 and 128 are in fluid communication with conduit 186 and are thus in fluid communication with each other. Conduit 186 may include valves 188 and 190 and pump 192. Conduit 186 is also in fluid communication with drain 194 and conduit 196. Conduit 196 is in fluid communication with reservoir 198. Reservoirs 104 and 198 may be connected to a means of sensing the mass thereof and as such may be connected to an electronic data processing network capable of receiving, transmitting, processing and recording data therefrom.

A re-circulation loop formed in the flow path includes conduit 128 (recirculation loop conduit) which can connect permeate outlets 170 and 172 via conduits 174, 176 and 186 (the permeate conduit) back to the retentate line/conduit (conduit 132) via conduits 118 and 125. This additional loop is a re-circulation loop that enables a system treatment solution (e.g., equilibration buffer/cleaning or storage solution) to circulate in the flow path until the user drains it via, for example, drain 194. Automated circulation through re-circulation loop minimizes the manual steps involved for filter and system conditioning as well as pre-filtration treatment and post-filtration treatment. Without the re-circulation loop, manual intervention would be needed when using a system treatment solution due to the additional connections/disconnections needed that would involve additional time and threaten user interference in the sterile condition of the system. Thus, utilizing the re-circulation loop, the sterility and/or integrity of the fluid system is not compromised as well as risk for compromising the sterility and/or integrity of the fluid system is greatly reduced by eliminating the manual interaction for connecting or disconnecting fluid conduits and containers at the outlet and/or inlet connections of the fluid system during the process. As a consequence, product (drug) and patient safety are increased. Further, operator safety during production of said products (drugs) is increased when processing potentially harmful fluids, fluid components or product intermediates, which could be the case during vaccine processing or production of ADC (antibody drug conjugates).

Some of the above components may be single use or multi-use depending on the system's usage. If components of the system are to be used more than once, the filter, the membrane therein and other system components can be cleaned using various washing fluid/buffers at the end of a filtration cycle to remove any contaminants (such as solids, particles, etc,) which may, for example, adsorb onto the membrane surface and block the pores. In this way, the operational lifetime of a filter, the membrane therein and other system components can be increased and their efficiency maintained.

The construction materials used in the system can suitably be compatible with commonly used sterilization methods, such as e.g. gamma irradiation and/or autoclaving. For reusable components, stainless steel (e.g. with corrosion resistance at least equivalent to 316 L) or engineering plastics such as polysulfone, PEEK, etc. may be used, while for single-use components, plastics, such as, e.g. polysulfone, polypropylene, polyethylene or ethylene copolymers, may be used. Tubing material may comprise materials such as silicone or TPE (thermoplastic elastomers); tubing may be weldable. All materials used in the construction of the system which come into contact with a solution (e.g., system treatment solution), biofluid, retentate and/or permeate are selected to avoid any chemical interaction and to minimize physical adsorption with the components within the solution. Typically, the valves are made of glass, ceramics, stainless steel or external tube pinch valves and the tubing of an inert plastic polymer.

All product contact surfaces of the system and components thereof are desirably, made of FDA compliant and/or USP Class VI tested materials. The system and its components should also be compatible with all commonly used solvents used in CFF/TFF, including, for example, IN NaOH (at 50° C.), 400 ppm NaOCl (at 50° C.), 1.1% phosphoric acid, 1.8% acetic acid, 2M HCl, 2M urea, “Triton-X” (a non-ionic detergent produced by polymerization of octylphenol with ethylene oxide, available from the Union Carbide Company, Danbury, Conn.), “Tween” (a polysorbate), 30-50% hexalene glycol, 30-50% propylene glycol, 0.07% polysorbate 20, 0.01-0.02% polysorbate 80, 90% ethanol, 90% methanol, 90% isopropyl alcohol, and 25% acetonitrile (w/v water).

To avoid contamination of the biofluid, all system components in contact with the biofluid should be suitably sterilized before the filtration process starts. The system or parts of the system may be assembled and sterilized by autoclaving or radiation, or one or more components may be pre-sterilized and assembled in a sterile system. To facilitate assembly, the system parts or components may be equipped with and connected by suitable connectors, in particular, the sterilized system parts or components may be equipped with and connected by aseptic connectors, e.g. of the ReadyMate type (GE Healthcare). As such, the sterilized system parts or components and associated connectors may be packaged in aseptic packaging. The sterilized system parts/components may also be contained in aseptic packages and assembled in a controlled environment, e.g., a clean room.

Typical filling operation of an automated CFF/TFF system is exemplified in FIG. 2. Although the solution to be filtered, for example, a biofluid, is contained during operation in reservoir 200, in typical practice, the reservoir 200 may not necessarily be the starting point or origin of the solution. For example, reservoir 200 can be provided with the solution from reservoir 202 through the conduit system shown in FIG. 2 driven by pump 204 with valves 206 in an open position and valves 208 in a closed position with the direction of flow indicated by arrows 210.

Typical filtration or concentration operation of the CFF/TFF system is exemplified in FIG. 3. The solution to be filtered is contained in reservoir 300 and proceeds through the conduit system shown in FIG. 3 through filter 302 driven by pump 304 with valves 306 in an open position and valves 308 in a closed position with the direction of flow indicated by arrows 310 for the feed, 312 for the retentate and 314 for the permeate. Valve 316 is open and valve 318 is closed when the permeate flow is to be sent to drain 320. Valve 316 is closed and valve 318 is open when the permeate flow is to be sent to reservoir 322. For example, if the solution includes a molecule of interest that is larger than the pore size of the filter membrane, the molecule will be present in the retentate that will be returned to reservoir 300 resulting in a greater concentration of biomolecule present in reservoir 300 as the operation proceeds. If, for example, the solution includes a molecule of interest that is smaller than the pore size of the filter, the molecule will be present in the permeate for collection, for example, in reservoir 322.

Prior to proceeding with the filtration operation, the system may need to treated with equilibration buffer or other pre-filtration treatment solutions to prepare the system for the filtration process. One of the purposes of equilibration buffer and other pretreating solutions may be to prepare the filter and membrane included therein as well as other system components for the fluid containing, for example, a desire biological product (e.g., a protein) by, for example, raising the salt concentration and other fluid physical parameters of the system including the filter, the membrane and other system components to match that of, for example, a bacterial lysate that will be put through the system.

Subsequent to the filtration process, if components of the system are to be used more than once, the filter, the membrane therein and other system components can be cleaned using various washing fluid/buffers at the end of a filtration cycle to remove any contaminants (such as solids, particles, etc,) which may adsorb to the membrane surface and block the pores.

Buffer equilibration and use of other pre-filtration treatment solution as well as use of post filtration washing fluid/buffers and storage solutions (all of which are referred to herein as system treatment solutions) is accomplished by the exemplified embodiment as shown in FIGS. 4, 5 and 6. In FIG. 4, the exemplary embodiment undergoes the step of system treatment solution fill in which reservoir 400 may provided with the system treatment solution from reservoir 402 through the conduit system shown in FIG. 4 driven by pump 404 with valves 406 in an open position and valves 408 in a closed position with the direction of flow indicated by arrows 410.

In FIG. 5, the system treatment solution from reservoir 500 is circulated through the conduit system shown in FIG. 5 and filter 502 driven by pump 504 with valves 506 in an open position and valves 508 in a closed position with the direction of flow indicated by arrows 510. Re-circulation is important to the filtration process. For example, buffer needs to recirculate through the filter for filter conditioning at a particular flow for stipulated time and in the same way for the filter cleaning and storage. Conditioning will improve filter efficiency. Suitable pump flow rates during this process may range from about 0.1 liter per hour to about 5000 liters per hour, preferably about 1 liter per hour to about 1000 liters per hour, more preferably 2-300 liters per hour. The conditioning circulation process can take place from about 10 minutes to about 6 hours, however, the time can vary depending on such parameters as, for example, filtration area, filter membrane type, filter condition and process condition.

In FIG. 6, after system treatment solution circulation is completed, the system treatment solution is drained from reservoir 600, filter 602 and the conduit system. The draining process is driven by pump 604 with valves 606 in an open position and valves 608 in a closed position with the direction of flow indicated by arrows 610 and to drain 612.

In at least one aspect of the disclosed embodiments, the systems and methods disclosed herein may be executed by an electronic data processing network including, for example, one or more controllers, computers or processor-based components under the control of one or more programs stored on computer readable medium, such as a non-transitory computer readable medium. FIG. 7 shows a block diagram of an exemplary controller or computing apparatus 700 that may be used to practice aspects of the disclosed embodiments. In at least one exemplary aspect, the system control circuitry, data acquisition circuitry, data processing circuitry, operator workstation and other disclosed devices, components and systems may be implemented using an instance or replica of the controller or computing apparatus 700 or may be combined or distributed among any number of instances or replicas of computing apparatus 700.

The controller or computing apparatus 700 may include computer readable program code or machine readable executable instructions stored on at least one computer readable medium 702, which when executed, are configured to carry out and execute the processes and methods described herein, including all or part of the embodiments of the present disclosure. The computer readable medium 702 may be a memory of the controller or computing apparatus 700. In alternate aspects, the computer readable program code may be stored in a memory external to, or remote from, the apparatus 700. The memory may include magnetic media, semiconductor media, optical media, or any media which may be readable and executable by a computer.

Controller or computing apparatus 700 may also include a processor 704 for executing the computer readable program code stored on the at least one computer readable medium 702. In at least one aspect, computing apparatus may include one or more input or output devices to allow communication among the components of the exemplary imaging system, including, for example, what may be generally referred to as a user interface 706, such as, the operator workstation described above, which may operate the other components included in the imaging system or to provide input or output from the controller or computing apparatus 700 to or from other components of the imaging system. The controller or computing apparatus 700 may be connected to the pumps, valves, sensors and other components of the system (as illustrated in FIG. 9) for coordinating the operation thereof as well as receiving, transmitting, processing, and recording data from the pumps, valves, and sensors.

Preferably, the embodiment of FIG. 1 is a single use system that maximizes maintaining the sterility or integrity of the system during system operation such that the environment outside the system does not contaminate or violate the environment inside the system as well as vice versa. Such operation includes the process of treating the system with system treatment solutions (e.g., buffer, particularly equilibration buffer) and the filtration process itself. In order to maintain the integrity of the system, the embodiment may include a cabinet having sides that define a cabinet interior and cabinet exterior. Some of the components shown in FIG. 1 may be single use and mounted to the cabinet and may be partially exposed to an external side of the cabinet so as to provide easy access for installation and removal. As a result of utilizing single use components, the system can be set-up and system operation conducted automatically (e.g., utilizing electronic data processing network) without having to change or alter the path of system components that could affect the integrity of the system from the outside or allow components of solutions in the system (e.g., viruses, proteins and other components of a bio fluid) to escape the system that could, for example, potentially cause harm to human operators.

Preferably, the complete flow path in fluid contact is provided as a single use flow path unit that is exchangeable in between different processes and production runs. Single-use technology helps reduce time-consuming device preparations, such as cleaning and cleaning validation, and helps reduce a risk for cross-contamination, which is important in applications such as vaccine manufacturing, for example. For example, GE Healthcare Life Science's READYTOPROCESS™ platform provides disposable, scalable fluid processing solutions. For example, the ÄKTA ready™ single use chromatography system provides an integrated disposable fluid processing conduit system (Flow Kit) with tubing, sensors, pumps and other fluid treatment, control and monitoring components that comprises all system components in fluid contact aimed for exchange and disposal after processing with biofluids. The Flow Kit is manufactured in a controlled environment and subjected to a bioburden reduction by gamma irradiation.

External to the cabinet, for example, in FIG. 1 and the frontal 101, left 103 and right 105 side panels of the cabinet as shown in FIGS. 8 and 9 (back side panel not shown) may be reservoirs, conduit tubing, tubing connectors (e.g., aseptic connectors, such as the ReadyMate type (GE Healthcare)), sensors and intersection points to which the conduit tubing encounters other system components, for example pumps and a filter module. For example, some, if not all, of the conduit tubing is flexible tubing, preferably, single use tubing, (e.g., conduits 108, 110, 114, 118, 128, 132, 148, 174, 176, 186 and 196 in FIG. 1) external to the cabinet, including some flexible tubing mounted to an exterior side of the cabinet and other components such as valves, sensors and pumps positioned thereon. The reservoirs, preferably, single use reservoirs, (e.g., reservoirs 102, 104 and 198 in FIG. 1) may be mounted to the cabinet or separate therefrom. The CFF/TFF filter, preferably, a single use CFF/TFF filter, (e.g., filter 106 in FIG. 1) may be mounted to the cabinet such that the portion of the filter connected to the flexible tubing is external to the cabinet, for example, positioned on an external side of the cabinet. The sensors, preferably, single use sensors, (e.g., sensors 142, 144, 146, 158, 180, 182 and 184 in FIGS. 1, 8 and 9) may be mounted to the cabinet such that the portion of the sensor connected to the flexible tubing is external to the cabinet, for example, positioned on an external side of the cabinet. The valves (e.g., valves 112, 116, 122, 124, 126, 130, 136, 138, 140, 154, 156, 160, 178, 188 and 190 in FIGS. 1, 8 and 9) may be pinch valves mounted to the cabinet such that the portion of the valve through which the flexible tubing passes is external to the cabinet, for example, positioned on an external side of the cabinet. The pumps (e.g., pumps 120, 152 and 192 in FIGS. 1, 8 and 9) may be peristaltic pumps mounted to the cabinet such that the portion of the peristaltic pump through which the flexible tubing passes is external to the cabinet, for example, positioned on an external side of the cabinet. If another type of pump is utilized (e.g., pump 152 of FIG. 1), for example, a piston pump, the components of the pump that will encounter the solution passing through the system (e.g., inlet, outlet, chamber and parts contained therein, check valve, etc.) may be single use or replaced for each use as well. Furthermore with a piston pump, it may be mounted to the cabinet such that the portion of the piston pump connected to the flexible tubing is external to the cabinet, for example, positioned on an external side of the cabinet. An electronic data processing network including a controller or computing apparatus 700 connected to the pumps, valves, sensors and filter and capable of receiving, transmitting, processing and recording data related to as well as coordinating the operation thereof may be partially or completely internal to the cabinet.

As a result, the operations of system treatment with a system treatment solution and filtration of a solution (e.g., a biofluid) can be accomplished in succession without having to re-configure the system (such as re-routing the fluid conduits) that could compromise the integrity of the system.

Furthermore, an electronic data processing network can direct the other components of the system (e.g., pumps and valves as well as sensors and filter) to proceed with the operations of system treatment and filtration of a solution automatically and without human intervention.

This written description uses examples as part of the disclosure, including the best mode, and also to enable any person skilled in the art to practice the disclosed implementations, including making and using any devices or systems and performing any incorporated methods. The patentable scope is defined by the claims, and may include other examples that occur to those skilled in the art. Such other examples are intended to be within the scope of the claims if they have structural elements that do not differ from the literal language of the claims, or if they include equivalent structural elements with insubstantial differences from the literal languages of the claims. 

1. An automated CFF/TFF system configured to utilize flexible tubing, comprising a cabinet having sides that define a cabinet interior and a cabinet exterior, the cabinet including: a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior, and a plurality of sensor connectors such that when a sensor is connected thereto the portion of the sensor to which the flexible tubing is connected is positioned on the cabinet exterior; and an electronic data processing network at least partially positioned in the cabinet interior, the electronic data processing network connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors, wherein the plurality of pumps, the plurality of pinch valves and the plurality of sensor connectors are proximate to a path for the flexible tubing configured to form a fluid circuitry including a reservoir and a CFF/TFF filtration module through which a fluid stream of a solution may be conducted through the automated CFF/TFF system, the fluid circuitry comprising: a feed conduit to provide fluid communication between an outlet of the reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module, and a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit.
 2. The automated CFF/TFF system of claim 1, wherein the plurality of pumps and the plurality of valves are capable of driving and regulating the fluid stream through the automated CFF/TFF system.
 3. The automated CFF/TFF system of claim 1, wherein the plurality of sensor connectors when connected to a sensor are capable of acquiring data about the fluid stream as it flows through the automated CFF/TFF system.
 4. The automated CFF/TFF system of claim 1, wherein the electronic data processing network is capable of connecting to and determining the performance of the CFF/TFF filtration module.
 5. The automated CFF/TFF system of claim 1, wherein the fluid circuitry further includes a second reservoir suitable for containing the solution and having a second reservoir outlet that is in fluid communication with the reservoir.
 6. The automated CFF/TFF system of claim 1, wherein the plurality of pumps includes a peristaltic pump or a piston pump.
 7. The automated CFF/TFF system of claim 6, wherein the piston pump is single use.
 8. The automated CFF/TFF system of claim 1, wherein the flexible tubing, the sensors, the reservoir and the CFF/TFF filtration module are single use.
 9. A method of using an automated CFF/TFF system, the method comprising: providing an automated CFF/TFF system configured to utilize flexible tubing comprising: a cabinet having sides that define a cabinet interior and a cabinet exterior, the cabinet including: a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior, a plurality of sensor connectors such that when a sensor is connected thereto the portion of the sensor through which the flexible tubing is connected is positioned on the cabinet exterior; and an electronic data processing network at least partially positioned in the cabinet interior, the electronic data processing network connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensors, wherein the plurality of pumps, the plurality of pinch valves and the plurality of sensors are proximate to a path for flexible tubing configured to form a fluid circuitry including a first reservoir, a second reservoir, a third reservoir and a CFF/TFF filtration module through which a fluid stream of a solution may be conducted through the automated CFF/TFF system, the fluid circuitry comprising: a feed conduit to provide fluid communication between an outlet of the first reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the first reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module, a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit, a second reservoir feed conduit to provide fluid communication between the first reservoir and the second reservoir, a third reservoir feed conduit to provide fluid communication between the first reservoir and the third reservoir, and a drain; connecting a sensor to at least one of the plurality of sensor connectors; forming the fluid circuitry using flexible tubing to connect the first reservoir, the second reservoir, the third reservoir, the CFF/TFF filtration module, the plurality of pinch valves, the plurality of pumps and the plurality of sensors; configuring the plurality of pumps and plurality of valves to provide a system treatment solution to the first reservoir from the second reservoir; providing a system treatment solution to the first reservoir from the second reservoir; configuring the plurality of pumps and plurality of valves to circulate the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; circulating the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; configuring the plurality of pumps and plurality of valves to drain the system treatment solution from the automated CFF/TFF system using the drain; draining the system treatment solution from the automated CFF/TFF system using the drain; configuring the plurality of pumps and plurality of valves to provide a solution for filtration to the first reservoir from the third reservoir; providing a solution for filtration to the first reservoir from the third reservoir; configuring the plurality of pumps and plurality of valves to filter the solution for filtration from the first reservoir through the CFF/TFF filtration module; and filtering the solution for filtration from the first reservoir through the CFF/TFF filtration module.
 10. The method of claim 9, further including driving and regulating the fluid stream through the automated CFF/TFF system using the plurality of pumps and the plurality of valves.
 11. The method of claim 9, further including acquiring data about the fluid stream through the automated CFF/TFF system using the plurality of sensors.
 12. The method of claim 9, wherein the CFF/TFF filtration module includes sensors capable of determining the performance thereof, connecting the electronic data processing network to the CFF/TFF filtration module so as to at least one of receiving, transmitting, processing and recording data associated with CFF/TFF filtration module and determining the performance of the CFF/TFF filtration module using the electronic data processing network.
 13. The method of claim 9, wherein the drain is in fluid communication with the permeate conduit and the recirculation loop conduit; and draining the system treatment solution includes circulating the system treatment solution from the first reservoir through the feed conduit, the automated CFF/TFF filtration module, the retentate conduit, the permeate conduit, the recirculation loop conduit to the drain such that the flow in the retentate conduit proceeds to the recirculation loop conduit and the flow from the recirculation loop conduit and the flow from the permeate conduit exit the automated CFF/TFF system through the drain.
 14. The method of claim 9, wherein the system treatment solution includes buffer.
 15. The method of claim 9, wherein the solution for filtration includes a biofluid.
 16. The method of claim 9, wherein the electronic data processing network controls the steps of: configuring the plurality of pumps and plurality of valves to provide a system treatment solution to the first reservoir from the second reservoir; providing a system treatment solution to the first reservoir from the second reservoir; configuring the plurality of pumps and plurality of valves to circulate the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; circulating the system treatment solution from the first reservoir through the feed conduit, the CFF/TFF filtration module, the retentate conduit, the permeate conduit and the recirculation loop conduit to return to the first reservoir; configuring the plurality of pumps and plurality of valves to drain the system treatment solution from the automated CFF/TFF system using the drain; draining the system treatment solution from the automated CFF/TFF system using the drain; configuring the plurality of pumps and plurality of valves to provide a solution for filtration to the first reservoir from the third reservoir; providing a solution for filtration to the first reservoir from the third reservoir; configuring the plurality of pumps and plurality of valves to filter the solution for filtration from the first reservoir through the CFF/TFF filtration module; and filtering the solution for filtration from the first reservoir through the CFF/TFF filtration module, wherein the fluid circuitry remains unchanged during the above steps.
 17. The method of claim 9, wherein at least one of the plurality of pumps is a piston pump and the piston pump, the flexible tubing, the plurality of sensors, the first reservoir, the second reservoir, the third reservoir and CFF/TFF filtration module are single use.
 18. An automated CFF/TFF system, comprising a cabinet having sides that define a cabinet interior and a cabinet exterior, the cabinet including: a plurality of pinch valves such that the portion of the valve through which the flexible tubing passes is positioned on the cabinet exterior, a plurality of pumps such that the portion of the pump through which the flexible tubing passes or is connected is positioned on the cabinet exterior, and a plurality of sensor connectors including a sensor connected to at least one of the plurality of sensor connectors, the portion of the sensor to which the flexible tubing is connected is positioned on the cabinet exterior; an electronic data processing network at least partially positioned in the cabinet interior, the electronic data processing network connected to and capable of at least one of receiving, transmitting, processing and recording data associated with at least one of the plurality of pumps, the plurality of pinch valves and the plurality of sensors; and flexible tubing forming a fluid circuitry including a reservoir and a CFF/TFF filtration module through which a fluid stream of a solution may be conducted through the automated CFF/TFF system, the fluid circuitry comprising: a feed conduit to provide fluid communication between an outlet of the reservoir and a feed inlet of the CFF/TFF filtration module, a retentate conduit to provide fluid communication between a retentate outlet of the CFF/TFF filtration module and an inlet of the reservoir, a permeate conduit to provide fluid communication with a permeate outlet of the CFF/TFF filtration module, and a re-circulation loop conduit to provide fluid communication between the permeate conduit and the retentate conduit.
 19. The automated CFF/TFF system of claim 18, wherein at least one of the plurality of pumps is a piston pump and the piston pump, the flexible tubing, the plurality of sensors, the reservoir and CFF/TFF filtration module are single use.
 20. The automated CFF/TFF system of claim 18, wherein the electronic data processing network is connected to and capable of determining the performance of the CFF/TFF filtration module. 